Vitrification of donkey sperm using straws as an alternative to conventional slow freezing

Cryoprotectant-free vitrification of donkey sperm using 0.25 ml straws has been recently developed, but the obtained results have not been directly compared to conventional slow freezing yet. The aim of this study was to compare sperm quality parameters after cryopreservation using both methods. Semen samples were collected from three Andalusian Donkeys. Semen was centrifuged and pellets resuspended with an extender with glycerol for conventional freezing or the same extender without glycerol, but with sucrose 0.1 mol/L for vitrification. Conventional freezing was performed in nitrogen vapours and thawed in a water bath (30s/37°C). Vitrification was performed in covered 0.25 ml straws plunged directly into liquid nitrogen and warmed in 3 ml of a milk-based extender at 43°C. Total (TM, %) and progressive motility (PM, %) were evaluated by computer-assisted sperm analysis, and plasma membrane (PMI, %) and acrosome (AIS, %) integrities by epifluorescence microscopy. No differences (p > .05) were found between slow freezing and vitrification methods for any of the parameters assessed: TM (58.2 ± 16.1% vs. 52.7 ± 15.6%), PM (44.7 ± 18.2% vs. 44.3 ± 15.0%), PMI (55.4 ± 9.0% vs. 49.2 ± 11.2%) and AIS (38.4 ± 19.6% vs. 45.0 ± 11.0%), respectively. In conclusion, donkey sperm vitrification in straws presented similar sperm quality after thawing in comparison to conventional freezing. Therefore, it could be considered as an alternative to slow freezing regarding the sperm parameters assessed.